The invention relates generally to the identification of agents for treating pathophysiologies associated with peroxisomal proliferator activated receptor-xcex1 (PPARxcex1), and of identifying ligands for PPARxcex1, and of identifying agents for treating these pathophysiologies. More particularly, the invention relates to methods of diagnosing these pathophysiologies, and of identifying agents for treating these pathophysiologies, using nucleic acids whose expression changes following addition to cells of a ligand for PPARxcex1.
Elevated low-density lipoprotein (LDL) cholesterol has long been recognized as an independent risk factor for the development of coronary artery disease and subsequent myocardial ischemia and infarction. There is also growing evidence that elevated triglycerides may also present a significant independent risk factor for development of coronary heart disease.
Increased triglyceride-rich lipoprotein remnants play a central role in increased atherosclerosis in several dyslipoproteinemias. Elevated triglycerides associated with low HDL is a frequent genetic dyslipoproteinemia in patients with established cardiovascular disease.
The fibrate class of lipid lowering agents have been demonstrated effective in managing coronary artery disease. Fibrates include the marketed drugs clofibrate (Atromid), Gemfibrozil (Lopid), Fenofibrate (Lipidil), ciprofibrate (Lipanor) and the experimental compound bezafibrate, These fibrates have been characterized as peroxisome proliferators based upon their ability to increase peroxisome number and activity in rodent model systems. Molecular characterization of the peroxisome proliferator activated receptor xcex1 (PPARxcex1), and subsequent pharmacological analysis of the fibrate compounds has demonstrated that these compounds are ligands of the PPARxcex1. PPARxcex1 is expressed in the liver, kidney and heart. Natural ligands for the receptor include C16-C20 polyunsaturated monocarboxylic fatty acids (PUFA).
The lipid lowering effects of fibrates are directly mediated through PPARxcex1. These receptors are members of the nuclear receptor superfamily of ligand-dependent transcription factors. Three subtypes of mammalian PPAR""s have been characterized: -alpha, -gamma, -delta. Members of the PPAR family exert their effect on transcriptional regulation through heterodimerization of ligand-bound PPAR receptors with the retinoid X receptor (RXR). These activated receptors bind to PPAR response elements (PPRE""s) in DNA to initiate a transcriptional response.
Peroxisomes, a class of subecclular organelles with which PPARxcex1 is associated, are enclosed by a single-layered membrane. Currently, more than 50 metabolically relevant enzymatic activities have been characterized within the peroxisome. These include xcex2-oxidation of long-chain fatty acids and derivatives (dicarboxylic acids, prostanoids, some xenobiotics and the side chain of cholesterol), fatty acid elongation, the hydrolysis of acyl-CoA""s and their conversion to acylcamitines, biosynthesis of ether glycerolipids, cholesterol and dolichols, the catabolism of purines, polyamines and amino acids and the metabolism of reactive oxygen species. Various in vivo and in vitro drug-dosing models have each associated one or two genes from these pathways with PPARxcex1 ligand activity. However, a comprehensive analysis of PPARxcex1 ligand-mediated activity has not been described.
The present invention is based in part on the discovery of changes in expression patterns of multiple nucleic acid sequences in rodent liver cells following exposure to a ligand for a PPARxcex1. The nucleic acid sequences whose expression changes include nucleic acid sequences encoding previously described polypeptides, as well as heretofore undescribed nucleic acid sequences. Expression of a subset of the sequences increased in liver cells following addition of the PPARxcex1 ligand, while expression of other sequences decreased following exposure to the ligand.
Based in part on the identification of these PPARxcex1 ligand responsive nucleic acid sequences, the invention provides screening methods based on nucleic acid sequences responsive to PPARxcex1 ligands. Also provided are methods for diagnosing or assessing conditions associated with PPARxcex1 metabolism using genes differentially expressed in response to PPARxcex1 ligands, as well as methods of treating adrenoleukodystrophy using PPARxcex1 ligands. In other aspects, the invention provides nucleic acid collections for identifying agents and pathologies associated with PPARxcex1 ligand responsive nucleic acid sequences. The invention additionally provides substrate arrays for identifying agents and pathologies associated with PPARxcex1 ligand responsive nucleic acid sequences, as well as single nucleotide polymorphisms associated with PPARxcex1 ligand responsive genes. The PPARxcex1-mediated disorders described herein can include, e.g, the pathophysiology is selected from the group consisting of: adrenoleukodystrophy; hyperlipidemia; peroxisomal disorders; dyslipidemia; hypertriglyceridemia; coronary artery disease; myocardial ischemia and infarction; and disorders associated with lipid metabolism, fatty acid metabolism, ketogenesis, microsomal-oxidation, and fatty acid-oxidation.
Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Although methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present invention, suitable methods and materials are described below. All publications, patent applications, patents, and other references mentioned herein are incorporated by reference in their entirety. In the case of conflict, the present specification, including definitions, will control. In addition, the materials, methods, and examples are illustrative only and not intended to be limiting.
Other features and advantages of the invention will be apparent from the following detailed description and claims.